Bender JK, Rydzewski K, Broich M, Schunder E, Heuner K, Flieger A (2009): Phospholipase PlaB of Legionella pneumophila represents a novel lipase family: protein residues essential for lipolytic activity, substrate specificity, and hemolysis
J. Biol. Chem. 284 (40): 27185-27194. Epub Jul 29.

Legionella pneumophila possesses several phospholipases capable for host-cell manipulation and lung damage. Recently, we discovered the major cell-associated hemolytic phospholipase A PlaB sharing no homology to described phospholipases and being dispensable for intracellular replication in vitro. Nevertheless, here we show that PlaB is the major lipolytic activity in L. pneumophila cell infections and that PlaB utilizes a typical catalytic triad of Ser-Asp-His for effective hydrolysis of phospholipid substrates. Crucial residues were found to be located within the N-terminal half of the protein and amino acids embedding these active sites were unique for PlaB and homologs. We further displayed that catalytic activity towards phosphatidylcholine, but not phosphatidylglycerol is directly linked to hemolytic potential of PlaB. While the function of the prolonged PlaB C-terminus remains to be elucidated, it is essential for lipolysis as removal of fifteen amino acids already abolishes enzyme activity. Additionally, we determined PlaB to preferentially hydrolyse long-chain fatty acid substrates containing 12 or more carbon atoms. As phospholipases play an important role as bacterial virulence factors, we examined cell-associated enzymatic activities among L. pneumophila clinical isolates and non-pneumophila species. All tested clinical isolates showed comparable, while out of the non-pneumophila species, only L. gormanii and L. spiritensis possessed similar lipolytic activities as L. pneumophila and comprised plaB-like genes. Interestingly, phosphatidylcholine-specific phospholipase A activity and hemolytic potential was more pronounced in L. pneumophila. Therefore, hydrolysis of the eukaryotic membrane constituent phosphatidylcholine triggered by PlaB could be an important virulence tool for Legionella pathogenicity.